The objective of this project is to identify and define structural features of terminal complement proteins whichmediate their interactions with each other and with the cell membrane during the cytolytic process. Proposed studies will focus specifically on structure-function relationships in the eight C8) and ninth (C9) components of human complement. Initial experiments will be concerned with characterizing structural domains of C8 whch are recognized by C5b-7, the membrane-bound precursor of C5b-8. Because such domains reside in the Beta subunit of C8, experiments will focus on this particular subunit andon identifying structural features which mediate its interaction with the C8 binding site on C5b-7. Other experiments in this project will examine the role of the Alpha-Gamma and Beta subunits of C8 in mediating C9 interaction with C5b-8, the precursor of the C5b-9 lytic complex. This will be accomplished through binding studies and the use of intrinsic cross-linking reagents. The latter will be used to establish the topographical relationship between Alpha-Gamma, Beta and C9 and to examine their proximity to other constituents in the terminal complexes. The role of Alpha-Gamma, Beta and C9 in membrane lysis willalso be examined. Here, protein components of membrane-bound C5b-8 and C5b-9 which insert into thelipid bilayer during lysis will be identified using a lipophilic, radiolabeled photosensitive reagent incorporated into membranes or phospholipid vesicles. The intramembrane peptide regions of Alpha, Beta, Gamma and C9 will be isolated and characterized with regard to amino acid composition and sequence. Use of this approach, along with concurrent determination of the complete sequence of C8 and C9, will facilitate the identification of localized hydrophobic sequences within these proteins and should provide conclusive evidence that such domains mediate the cytolytic function of C5b-9 by insertion into the bilayer. When completed, this project will contribute to understanding the molecular basis of terminal complement protein function. Because it is concerned with understanding the mechanism and regulation of an intrinsic, physiological process for destroying cells, it also has substantial biological significance with regard to both infectious diseases and potential improvements in the treatment of cancer.